摘要
从广西部分地区采集到自然罹病死亡的疑似具有病毒症状的斜纹夜蛾幼虫,对其进行病毒的分离鉴定。用研磨病虫尸体得到的浆液,经分离纯化后感染连续人工饲养至第三代的健康斜纹夜蛾三龄幼虫,部分样品能诱发病毒病症状。经差速离心的提纯物,在相差显微镜下观察到外形不规则的颗粒,进一步在电镜下观察到菱形、五边型、六边形、近似圆形等形状的多角体。用依据Splt NPV DNA的egt基因保守序列设计的引物,对多角体中提取的DNA进行PCR扩增,得到了与预期一致的3 80 bp片段。经测序分析,该PCR片段与Splt NPV egt基因在核苷酸水平上的同源性为99.4%,演绎的氨基酸水平上的同源性为1 0 0 %。进一步对采自广西1 6个地域的疑似具有病毒症状的2 0 0份斜纹夜蛾幼虫样本进行PCR扩增,其中6个地域的62份样本呈PCR阳性,这表明,广西斜纹夜蛾自然种群中存在着Splt
Larval cadavers of Spodoptera litura that were likely to die from viral diseases were collected from regions of Guangxi province.Partially purified homogenate of the diseased larvae were used to feed healthy larvae of S.litura that had been propagated to 3 rd generation under controlled caged condition.Syndromes of typical S.litura nucleopolyhedrovirus ( Splt NPV) infection were observed in some of the samples. Purified precipitate from homogenate were examined under phase- contrastmicroscope,and irregularpolyhedrons were found.Polyhedrons of diamond,pentagon,and hexagon ornearrotundity were observed when the sampleswere examined underelectron microscope. Expected fragments of 3 80 bp could be amplified using PCR primers that were designed according to the conserved region of egt gene of Splt NPV when the DNA from the precipitates were used atemplates.The PCR productwas sequenced and blasted against Gene Bank database.Itwas revealed thatthe sequence had homologies of99.4% at nucleotide level and1 0 0 % atdeduced amino acid level to Splt NPV. Two hundred larval cadavers of S.litura from 1 6 regions of Guangxi province were further amplified by PCR,and6 2 samples from6 regions were positive.The results showed that Splt NPV was prevalentnaturally in population of S.litura in some regions of Guangxi province.
出处
《广西农业生物科学》
CAS
CSCD
2005年第1期9-13,共5页
Journal of Guangxi Agricultural and Biological Science
基金
微生物及植物遗传工程教育部重点实验室开放基金 ( J0 40 2 )
关键词
斜纹夜蛾
核型多角体病毒
分离鉴定
Spodoptera Litura
Nucleopolyhedrovirus
Isolation and identification
